Project Summary/Abstract A major focus of the UC San Diego Superfund Research Program Center (SRC) is on the impact of toxicant exposure on the development of liver cancers and fibrosis with an emphasis on Toxicant-Associated Steatoheptatitis (TASH). As part of this project, we focus on the role of several hazardous substances on the Priority List of Hazardous Substances by the Agency for Toxic Substances and Disease Registry (ATSDR). Current analytical techniques used for the detection of such pollutants/toxicants are generally expensive to use and typically done in the lab setting. Clearly, there is a great need for cheap biosensors for specific detection of toxicants that is: (1) inexpensive to produce, (2) easy to use, (3) portable/deployable in the field, and (4) a game- changing research tool for studying environmental health sciences. We have, therefore, developed an innovative technique that will make it possible to detect and quantitate metals and other environmental toxicants by creating genetically encodable molecular sensors. Using a new and powerful molecular evolution platform, we combine non-homologous site-to-site recombination on modest sized libraries with selection strategies to discover protein biosensors that light upon addition of a specific ligand or small molecular weight toxicant. These biosensors can also be made to become molecular switches that will turn ?ON? and light/fluoresce with desired color/wavelength upon binding as well as reversibly turn ?OFF? when the small molecules diffuse away, providing a dynamic measure of its concentration. Protein biosensors and switches will allow us to track toxicants such as metals (As, Cd, Pb, Hg) and other ubiquitous environmental agents such as organochlorides, polycylics, and dioxins. We envision developing panels of inexpensive small-molecule biosensors that can be spotted on filter paper, for example, to do rapid on-site detection of toxicants and metals.